Dosage compensation occurs to equalize X-linked gene expression in males (XY) and females (XX), and is essential in many organisms that utilize a chromosomal basis for sex determination. In Drosophila, dosage compensation occurs primarily by increasing transcription of X-linked genes in males. The genetic switch that confers sex-specificity is critical: lack of X chromosome hypertranscription kills males, while inappropriate X hypertranscription kills females. Recent studies suggest that dosage compensation requires a distinct chromatin environment on the male X. Four regulatory proteins (MSL proteins) and a specific acetylated isoform of histone H4 (H4Ac16) are associated in a banded pattern along the male X and not along the autosomes or the female X chromosomes. The presence of these proteins correlates with the distinct cytological appearance of the male X, and its increased transcriptional activity. Our experiments focus on the function and regulation of the MSL proteins. We will analyze the mechanism of dosage compensation as a model for the coordinate regulation of genes by their chromatin environment. We will also dissect the genetic switch by which dosage compensation is regulated. The specific aims are: I. Analysis of the link between site-specific histone acetyltransferase activity and dosage compensation. II. Identification of the cis-acting sites that confer X chromosome- specificity to the MSL proteins. III. Definition of the sex-specific mechanism that regulates dosage compensation. IV. Identification of additional dosage compensation regulators using novel genetic screens. V. Analysis of sex lethal action in MSL-independent dosage compensation. Both biochemical and genetic approaches will be utilized. The biochemical studies will focus on identification of a male-specific histone H4 acetyltransferase activity, and its relationship to the MSL proteins. Definition of chromatin targets of the MSL proteins will address the basis for their exquisite X chromosome specificity. Transgenic experiments will dissect the regulation and function of the previously cloned msl genes. Novel genetic screens will capitalize on transgenic phenotypes to identify new functions required for dosage compensation. Understanding how the MSL proteins function and are regulated is likely to relate to how genes are controlled during development in higher organisms. The MSL proteins act in the establishment and maintenance of functional states of chromatin, which is a subject at the frontier of research on transcriptional regulation. In addition, the sex-specific regulation of the MSL proteins provides a new model for a genetic switch. Bi-modal switch mechanisms found initially in model organisms such as fruitflies are in many cases relevant to normal and disease states in humans.